Project Summary/Abstract Virtually every aspect of human physiology and behavior is modulated by an inherent 24 hour (circadian) timing process. At the center of this clock timing system is the suprachiasmatic nucleus (SCN) of the hypothalamus. A key feature of the SCN clock is the tight, time-of-day, dependent regulation of the MAPK (p44/42 mitogen-activated protein kinase) pathway. Two examples of this phenomenon are the daily oscillations in the activation state of the MAPK pathway, and the clock-gated regulation of the photic responsiveness of the pathway. Importantly, the clock-generated, temporally-delimited, regulation of MAPK signaling appears to play a central role in SCN timing and entrainment. Further, the daily gating of MAPK signaling may be an underlying design principal of all oscillator populations, and as such, MAPK rhythms could have profound and far-reaching effects on a range of physiological processes. Given these implications, it is surprising that we still know relatively little about the cellular mechanisms and synaptic circuits that confer circadian control over MAPK activity. Here, we hypothesize that the circadian regulation of MAPK signaling is an inherent (cell autonomous) feature of SCN cellular oscillators and that this MAPK rhythm is a key mechanistic building-block by which the circadian clock modulates both basic and complex physiological states. To test this hypothesis, we propose the following set of experimental goals. In Aim 1, we will identify the cellular and network properties of the SCN that give rise to the rhythmic regulation of the MAPK pathway. To this end, we will, A) Determine whether MAPK rhythms are cell autonomous or whether they result from an intercellular SCN network, and B) Determine the intracellular signaling events that generate MAPK activity rhythms. In Aim 2 we propose to characterize the molecular, cellular and systems-based mechanisms by which the SCN clock gates light-evoked MAPK pathway activation. To address this largely unexplored phenomenon, we will, A) determine when and how the molecular gate opens, and B), test whether the cytoplasmic ERK scaffold protein PEA-15 serves as the principal circadian gate on MAPK signaling. Of note, we recently identified PEA-15 as a modulator of MAPK signaling in the SCN, and its capacity to dynamically regulate ERK signaling makes it an attractive candidate for the gating of MAPK signaling. In Aim 3 we propose to employ a selective targeting approach to transgenically disrupt MAPK signaling within the SCN core and shell regions to address the roles of MAPK signaling in A) the generation of circadian rhythms, and B) the entrainment of the circadian clock. Further, conditional PEA-15 KO and point mutant PEA-15 transgenic mouse lines will be used to test a model in which PEA-15 phosphorylation leads to rapid ERK dissociation, which we posit to be a key step in the initiation of light-evoked phase-shifting. Together, these data will provide fundamental new insights into the relationship between MAPK signaling and the circadian clock, and point to potential ways in which the dysregulation of clock-gated MAPK signaling could contribute to disorders of the CNS.